Nowadays, because of the lack of useful in vitro procedures, in order to obtain a polyclonal antibody preparation, one must immunize an animal (such as a rabbit, a goat, a mouse, a rat, etc.) with an antigen of interest and purify the antibodies generated from the animal. Some industries even specialize in this type of service. The resulting polyclonal antibody preparations are labor-intensive and costly.
Some systems have been developed for the large-scale in vitro production of monoclonal antibodies. One process widely used is the production of hybridomas (e.g., immortalized B-cell lines capable of secreting a specific monoclonal antibody). However, hybridomas are not necessarily stable and may lose their ability to proliferate or secrete immunoglobulins as they are cultured. Attempts have been made to produce more stable hybridomas. For example, U.S. application Ser. No. 11/509,364 (published under 2007/0098711 on May 3, 2007) describes a method for stabilizing an antibody-secreting cell by repeated oligoclonal handpicking.
Other culture systems have been developed to generate more stable B-cell cultures capable of proliferation and/or differentiation. One of these systems is the CD40L or CD154 system. For example, U.S. Pat. No. 5,817,516 (granted on Oct. 6, 1998) and U.S. Pat. No. 6,297,052 (granted on Oct. 2, 2001) describe the use of CD40L to facilitate the proliferation and the differentiation of B cells cultured in vitro. However, the isotypes of immunoglobulins (Ig) harvested under the conditions described in U.S. Pat. No. 5,817,516 or 6,297,052 are not similar to those observed in peripheral blood, suggesting that these methods introduce bias in the secretion of Ig.
The CD40-CD154 culture system is a model of the in vivo interaction between CD40 present on B cells and CD154 present on activated T cells1. The utility of this model has been demonstrated by Banchereau et al. in 1991, as it allows to grow human B cells in culture independently of classical mitogens and/or antigenic stimulation and to prepare human immunoglobulins and monoclonal antibodies2. At that time, Banchereau used B cells isolated from human tonsils3. Since then, many other investigators have used this culture system as a means of activating human B cells from other sources, such as the spleen4 and blood5-7, to study their physiological characteristics in relation to the immune response. Most groups have been sorting B cells on the basis of the expression of CD27 molecules on the cell surface, CD27+ cells being recognized as memory B cells, and CD27− cells as naïve B cells, respectively involved in secondary and primary immune responses.
It would be highly desirable to be provided with an in vitro method for the production of a large quantity of immunoglobulin G. This method should enable the proliferation and the differentiation of IgG-secreting B cells. Preferably, this method should allow the large-scale production of human IgGs.